Studies with reassortant viruses which contained human influenza A virus surface antigens from the A/Washington/80 (H3N2) virus and one or more internal genes from the avian A/Pintail/Alberta/79 virus demonstrated: (1) a particular constellation of polymerase genes specifies a host range dependent restriction of replication in MDCK but not chick kidney tissue culture, (2) the ability to replicate at 42 degrees C is a complex, multigene-associated phenotype, and (3) the NP gene is the major contributor to attenuation in monkeys. Comparison of the nucleotide and amino acid sequences of the NP genes of the avian A/Mallard/NY/78 with those of two other avian and three human influenza A viruses showed the existence of separate avian and human classes of NP genes with separate evolutionary patterns. The rate of evolution of human influenza A virus NP genes in the first several years immediately following the emergence of a new subtype in 1968 was increased compared with other years. Studies with single gene reassortant viruses derived from the human influenza A/Ann Arbor/6/60 ca virus confirmed the role of polymerase PA in specifying the ca phenotype and that of the polymerase PB2 in specifying the ts phenotype. The PA polymerase and M genes appear to play a role in attenuation. The mechanism of attenuation by the M gene is independent of ts or ca phenotypic markers. Reassortant viruses which contained a wild type influenza A/Alaska/77 virus NS gene which had sustained a 36 base deletion mutation in the region coding for the NS1 protein were studied. In association with "internal genes" from the ca donor virus, the mutant gene specified: (a) a host range restriction of replication in MDCK but not in chick kidney tissue culture, and (b) a ts phenotype. A reassortant containing the mutant NS in association with wild type genes exhibited only the ts phenotype. Loss of the ts phenotype occurred after replication of this reassortant in hamsters and chimpanzees.